Inhibition of phosphatase activity as a novel treatment for chronic toxoplasmosis

描述(申请人提供)：本申请针对由原生动物寄生虫弓形虫引起的潜伏性慢性感染。在宿主体内，弓形虫从导致急性感染的增殖期(速殖子)转变为潜伏的包囊(缓殖子)阶段，从而形成终生慢性感染。慢性弓形虫病具有非常重要的临床意义，因为潜伏感染可以重新激活为危及生命的急性弓形虫病，尤其是在艾滋病和心脏移植患者中。这项应用解决了开发能够防止潜伏囊肿重新激活感染的治疗方法的重大需求。我们之前的发现证实了eIF2(真核细胞起始因子-2)磷酸化在微生物潜伏期中的作用， 这促使我们测试eIF2去磷酸化的抑制剂是否可以在体外干扰潜在的弓形虫感染的重新激活。我们发现，其中一种药物，愈创木酚(Ganabenz，GA)，是一种有效的抗寄生虫剂，在体外也能抑制缓殖子包囊的重新激活。重要的是，GA是一种耐受性良好的FDA批准的药物，可以跨越血脑屏障，这些特性使其成为快速发展为治疗慢性弓形虫病的创新新疗法的杰出候选者。在这项应用的R21阶段，我们将在一个特征良好的慢性感染小鼠模型中测试GA的实用性，并对GA耐药寄生虫进行筛选，以评估耐药的可能性和机制。这些突变体将促进研究，以确定这种药物如何抑制寄生虫的增殖和感染的重新激活。进展到R33阶段的里程碑包括在体内证明GA对慢性弓形虫的活性，或者产生GA耐药突变体。如果达到了这两个里程碑中的一个或两个，这项研究的R33阶段将阐明GA使用互补的生化和遗传方法控制寄生虫复制和潜伏感染的能力的分子机制。

DESCRIPTION (provided by applicant): This application addresses latent chronic infection caused by the protozoan parasite Toxoplasma gondii. In the host, Toxoplasma converts from a proliferative stage (tachyzoite) that causes acute infection to a latent cyst (bradyzoite) stage tht establishes a lifelong chronic infection. Chronic toxoplasmosis is of great clinical importance because the latent infection can reactivate into life-threatening acute toxoplasmosis, most notably in AIDS and heart transplant patients. This application addresses the significant need to develop therapies that can prevent latent cysts from reactivating infection. Our previous findings established a role for eIF2¿ (eukaryotic initiation factor-2) phosphorylation in microbial latency, prompting us to test if inhibitors of eIF2¿ dephosphorylation could interfere with reactivation of latent Toxoplasma infection in vitro. We found that one such drug, guanabenz (GA), is a potent anti-parasitic agent that also inhibits the reactivation of bradyzoite cysts in vitro. Importantly,GA is a well-tolerated FDA-approved drug that crosses the blood-brain barrier, properties that make it an outstanding candidate for rapid advancement as an innovative new treatment for chronic toxoplasmosis. In the R21 phase of this application, we will test the utility of GA in a well-characterized mouse model of chronic infection, and conduct a screen for GA-resistant parasites in order to assess the likelihood and mechanism of drug resistance. These mutants will facilitate studies to define how this drug inhibits parasite proliferation and the reactivatio of infection. Milestones for advancement to the R33 phase include either a demonstration of GA activity against chronic Toxoplasma in vivo or the generation of GA-resistant mutants. If one or both of these milestones are achieved, the R33 phase of this study will elucidate the molecular mechanism underlying the ability of GA to control parasite replication and latent infection using complementary biochemical and genetic approaches.